Project Summary The prefrontal cortex (PFC) is the brain area most associated with higher cognition. The ?modulatory? neurotransmitters acetylcholine (ACh) and serotonin (5-HT) facilitate these cognitive processes by regulating information ?ow within cortical circuits, and dysregulation of neuromodulatory input to the cortex is directly implicated in a number of mental health disorders. Despite their clear clinical importance, little is known about the physiological impact of ACh and 5-HT on synaptic transmission and integration within cortical neuron subpopulations. My preliminary data have revealed a reciprocal relationship between ACh and 5-HT in controlling two distinct and non-overlapping cortical output channels. 5-HT selectively inhibits, while ACh preferentially enhances, the output of corticopontine (CPn) neurons projecting to the brainstem, while 5-HT selectively enhances corticocortical (CC) neurons that project to the cerebral cortex. This differential regulation of CPn and CC neurons by 5-HT and ACh suggests circuit-based mechanisms by which these transmitters may in?uence cognition and behavior. The long-term objective of this project is to de?ne how modulatory neurotransmitters regulate the output of the cerebral cortex through selective, cell-type-speci?c pre- and postsynaptic actions. The short-term objective proposed here is to evaluate the effect of ACh and 5-HT on synaptic transmission from de?ned cortical afferents, and to characterize the feedback circuitry from the cortex to the cholinergic center of the basal forebrain. Speci?c Aim 1 will test the hypothesis that ACh and 5-HT in?uence transmitter release from axon terminals in ways that complement their actions on cellular excitability. Using innovative labeling strategies to express channelrhodopsin in select cortical projection neuron subpopulations, I will determine the postsynaptic speci?city of cortical afferents, test presynaptic modulation of glutamate release from speci?c cortical afferents, and identify the presynaptic receptors responsible for modulation of synaptic transmission. Speci?c Aim 2 will test the hypothesis that cortical output to the basal forebrain acts as a negative feedback loop by reducing cholinergic input to the cortex. Using retrograde and viral vector-based neuronal labeling, optogenetics, immunohistochemistry, and electrophysiology, I will identify the population of cortical neurons innervating the basal forebrain, and their net in?uence on cholinergic circuitry. A growing appreciation of the diversity of cortical neurons, their selective connectivity, and their differential responsivity to neuromodulatory transmitters, is expanding our understanding of cortical circuit function. The proposed project will contribute to that understanding by evaluating the effect of ACh and 5-HT on cortical microcircuits. Findings from this project have the potential to provide insight into how these two neuromodulators facilitate cognitive function, and may identify fundamental and conserved mechanisms contributing to information ?ow in other brain circuits.